Abaxis VetScan HM5, Operator'S Manual

Page: 129 / 164

Operating Principles B-3
Proper counting (or differentiation) of cells requires passing of only one cell through the aperture at a
time. To help ensure this, the blood samples must be diluted, since cell concentrations are otherwise too
high.
In cases of cell counts beyond the linear range, an external predilution of the sample is recommended.
B.2.2 Three-Part Differential Method
To perform a three-part WBC differential count, the RBCs must first be lysed since RBCs are typically
1000 times more numerous in normal blood than are WBCs and would interfere with WBC counting if
left intact. Lysing also releases the hemoglobin stored in the RBCs for direct analysis in the solution.
Therefore, a hemolysing reagent (Lyse) is used to dissolve cellular membranes, thus destroying RBCs,
and creating a complex solution suitable for photometry of HGB and counting WBCs.
The following figure shows the changes in blood cell characteristics that occur during three-part differ-
ential hemolysis.
Although diluted blood is used, in cases of
extremely high concentrations (such as
leukemia) WBC density can be 100x higher
than normal, causing two or more cells to
pass through the aperture at a time,
generating one pulse instead of two (or
more). This is called coincidence, and results
in non-linear counting of cells. Flags
m
,
M
,
and
N
appear in this case.
The WBC linearity range is 100 x 10
9
cells/l.